In 5-hydroxytryptamine-containing neurones of the mammalian central nervous system, one of the regulatory mechanisms for rapidly increasing transmitter synthesis during impulse flow is enhancement of the initial, rate-limiting step in 5-HT synthesis, the conversion of tryptophan to 5-hydroxytryptophan by tryptophan hydroxylase. Recent in vitro experiments in this laboratory have shown that tryptophan hydroxylase becomes activated after depolarization of slices of rat brain stem in a potassium-enriched incubation medium. The purpose of this proposal is to determine whether the enzyme also becomes activated and/or stimulated as a result of nerve impulse flow in vivo, and, if so, whether this activation reflects the ongoing level of activity in the 5-HT neurones. Experimental approaches to analyze this problem will include electrical stimulation of 5-HT neurones in the midbrain raphe, electrolytic lesions of 5-HT perikarya in dorsal or median raphe to block nerve impulse flow, and the use of drugs in vivo and in vitro which have reproducible and well characterized effects on firing of 5-HT neurones. Enzyme will be prepared from brain regions which receive projections from the midbrain raphe and its activity and kinetic properties determined with a very sensitive assay utilizing high pressure liquid chromatography and fluorimetric detection.